Investigation of mercury metallothionein complexes in tissues of rat after oral intake of HgCl2

Mercury, a highly toxic metal found widely throughout the environment, is a potent inducer of metallothionein (MT) expression. The role of MTs in the detoxification of mercury after its oral intake in mammals is studied. After feeding rats with mercuric chloride by gastric gavage, the distribution of heavy metals in rat tissues was investigated by inductively coupled plasma mass spectrometry (ICP-MS). Extensive accumulation of mercury, copper and zinc in kidney and liver is observed. A homemade preparative size-exclusion chromatography (SEC) column (30cm x 1.9cm) packed with Sephadex G-75 (40-120 mum particle size) gel (Pharmacia) was used for the purification of NIT fractions in rat tissues. Preliminary results from SEC indicate that the mercury-binding MT levels in liver were much lower than in kidney. The NIT fractions were collected, desalted, and then separated by reversed-phase high-performance liquid chromatography (HPLC) with UV-Vis spectrometry, ICP-MS and electrospray ionization MS detection. One major and several minor peaks were observed in the HPLC chromatograms of the MT fraction for the kidney sample. UV absorption spectra indicate that MTs were found to bind with mercury. There were no significant mercury-binding MTs detected in the liver sample using UV detection. ICP-MS detection showed that mercury-binding MTs in kidney contained large amounts of mercury and copper but little zinc. Further characterization with ESI-MS showed that the major peak found in kidney contained Hg6Cu, Hg5Cu2-MT-2c and Hg-6-MT-2beta, Hg6Cu-MT-1gamma, Hg-7-MT-2alpha. However, distinction between copper and zinc could not be made based on current mass spectrometric analysis because of instrumental resolution limitations. Copyright (C) 2004 John Wiley Sons, Ltd.